Anatomy of a conformational transition of β-strand 6 in soybean β- amylase caused by substrate (or inhibitor) binding to the catalytical site

Abstract

A computational study of the five soybean β-amylase X-ray structures reported so far revealed a peculiar conformational transition after substrate (or inhibitor) binding, which affects a segment of the β-strand 6 (residues 341-343) in the (β/α)8 molecular scaffold. Backbone distortions that involve considerable changes in the φ and ψ angles were observed, as well as two sharp rotamer transitions for the Thr342 and Cys343 side chains. These changes caused the outermost CA-layer (at the C-terminal side of the barrel), which is involved in the catalysis, to shrink. Our observations strongly suggest that the 341FTC343 residue conformations in the free enzyme are not optimal for protein stability. Furthermore, as a result of conformational transitions in the ligand-binding process, there is a negative enthalpy change for these residues (- 27 and -34 kcal/mol, after substrate or inhibitor binding, respectively). These findings support the proposed 'stability-function' hypothesis for proteins that recognize a ligand (Shoichet BK, Baase WA, Kuroki R, Matthews BW. 1995. A relationship between protein stability and protein function. Proc Natl Acad Sci USA 92:452-456). They are also in good agreement with other experimental results in the literature that describe the role of the 341-343 segment in β-amylase activity. Site-directed mutagenesis focused on these residues could be useful for undertaking functional studies of β-amylase.